Morphological examination of the highly polarized protozoan parasite Toxoplasma gondii suggests that secretory traffic in this organism progresses from the endoplasmic reticulum to the Golgi apparatus using the nuclear envelope as an intermediate compartment. While the endoplasmic reticulum is predominantly located near the basal end of the parasite, the Golgi is invariably adjacent to the apical end of the nucleus, and the space between the Golgi and nuclear envelope is filled with numerous coatomer-coated vesicles. Staining with antiserum raised against recombinant T. gondii beta-COP confirms its association with the apical juxtanuclear region. Perturbation of protein secretion using brefeldin A, microtubule inhibitors or dithiothreitol disrupts the Golgi, causing swelling of the nuclear envelope, particularly at its basal end. Prolonged drug treatment leads to gross distention of the endoplasmic reticulum, filling the basal end of the parasite. Cloning and sequencing of the T. gondii homolog of the chaperonin protein BiP identifies the carboxy-terminal amino acid sequence HDEL as this organism's endoplasmic reticulum-retention signal. Appending the HDEL motif to a recombinant secretory protein (a chimera between the parasite's major surface protein fusion, P30, and the Green Fluorescent Protein) causes this secretory reporter to be retained intracellularly. P30-GFP-HDEL fluorescence was most intense within the nuclear envelope, particularly at the apical end. These data support a model of secretion in which protein traffic from the endoplasmic reticulum to Golgi occurs via the apical end of the nuclear envelope.

REFERENCES

Ajioka
J. W.
,
Boothroyd
J.C.
,
Brunk
B. P.
,
Hehl
A.
,
Hillier
L.
,
Manger
I. D.
,
Marra
M.
,
Overton
G. C.
,
Roos
D. S.
,
Wan
K. L.
,
Waterston
R.
,
Sibley
L. D.
(
1998
).
Gene discovery by EST sequencing in Toxoplasma gondii reveals sequences restricted to the Apicomplexa.
Genome Res
8
,
18
28
Bangs
J. D.
,
Brouch
E. M.
,
Ransom
D. M.
,
Roggy
J. L.
(
1996
).
A soluble secretory reporter system in Trypanosoma brucei. Studies on endoplasmic reticulum targeting.
J. Biol. Chem
271
,
18387
18393
Barlowe
C.
,
Orci
L.
,
Yeung
T.
,
Hosobuchi
M.
,
Hamamoto
S.
,
Salama
N.
,
Rexach
M. F.
,
Ravazzola
M.
,
Amherdt
M.
,
Schekman
R.
(
1994
).
COPII: A membrane coat formed by Sec proteins that drive vesicle budding from the endoplasmic reticulum.
Cell
77
,
895
907
Becker
B.
,
Melkonian
M.
(
1996
).
The secretory pathway of protists: Spatial and functional organization and evolution.
Microbiol. Rev.
60
,
697
721
Bednarek
S. Y.
,
Ravazzola
M.
,
Hosobuchi
M.
,
Amherdt
M.
,
Perrelet
A.
,
Schekman
R.
,
Orci
L.
(
1995
).
COPI-and COPII-coated vesicles bud directly from the endoplasmic reticulum in yeast.
Cell
83
,
1183
1196
Braakman
I.
,
Hoover-Litty
H.
,
Wagner
K. R.
,
Helenius
A.
(
1991
).
Folding of influenza hemagglutinin in the endoplasmic reticulum.
J. Cell Biol
114
,
401
411
Burg
J. L.
,
Perelman
D.
,
Kasper
L. H.
,
Ware
P. L.
,
Boothroyd
J. C.
(
1988
).
Molecular analysis of the gene encoding the major surface antigen of Toxoplasma gondii.
J. Immunol
141
,
3584
3591
Carruthers
V. B.
,
Sibley
L. D.
(
1997
).
Sequential protein secretion from three distinct organelles of Toxoplasma gondii accompanies invasion of human fibroblasts.
Eur. J. Cell. Biol
73
,
114
123
Cesbron-Delauw
M. F.
,
Lecordier
L.
,
Mercier
C.
(
1996
).
Role of secretory dense granule organelles in the pathogenesis of toxoplasmosis.
Curr. Top. Microbiol. Immunol
219
,
72
65
Dieckmann-Schuppert
A.
,
Bause
E.
,
Schwartz
R. T.
(
1994
).
Glycosylation reactions in Plasmodium falciparum, Toxoplasma gondii, and Trypanosoma brucei brucei probed by the use of synthetic peptides.
Biochim. Biophys. Acta
1199
,
37
44
Duden
R.
,
Griffiths
G.
,
Frank
R.
,
Argos
P.
,
Kreis
T. E.
(
1991
).
-COP, a 110 kD protein associated with non-clathrin-coated vesicles and the Golgi complex, shows homology to -adaptin.
Cell
64
,
649
665
Dunn
P. P.
,
Bumstead
J. M.
,
Tomley
F. M.
(
1996
).
Primary structure of a BiP homolog in Eimeria spp.
Parasitol. Res
82
,
566
568
Ho
W. C.
,
Allan
V. J.
,
van Meer
G.
,
Berger
E. G.
,
Kreis
T. E.
(
1989
).
Reclustering of scattered Golgi elements occurs along microtubules.
Eur.J. Cell Biol
48
,
250
263
Hobman
T. C.
,
Zhao
B.
,
Chain
H.
,
Farquhar
M. G.
(
1998
).
Immunoisolation and characterization of a subdomain of the endoplasmic reticulum that concentrates proteins involved in COPII vesicle biogenesis.
Mol. Biol. Cell
9
,
1265
1278
Hong
W.
(
1998
).
Protein transport from the endoplasmic reticulum to the Golgi apparatus.
J. Cell Sci
111
,
2831
2839
Karsten
V.
,
Qi
H.
,
Beckers
C. J.
,
Reddy
A.
,
Dubremetz
J. F.
,
Webster
P.
,
Joiner
K. A.
(
1998
).
The protozoan parasite Toxoplasma gondii targets proteins to dense granules and the vacuolar space using both conserved and unusual mechanisms.
J. Cell Biol
141
,
1323
1333
Klausner
R. D.
,
Donaldson
J. G.
,
Lippincott-Schwartz
J.
(
1992
).
Brefeldin A: Insights into the control of membrane traffic and organelle structure.
J. Cell Biol
116
,
1071
1080
Köhler
S.
,
Delwiche
C. F.
,
Denny
P. W.
,
Tilney
L. G.
,
Webster
P.
,
Wilson
R. J. M.
,
Palmer
J. D.
,
Roos
D. S.
(
1997
).
A plastid of probable green algal origin in apicomplexan parasites.
Science
275
,
1485
1488
Kumar
N.
,
Zheng
H.
(
1992
).
The nucleotide sequence of a Plasmodium falciparum stress protein with similarities to a mammalian 78 kDa glucose-regulated protein.
Mol. Biochem. Parasitol
56
,
353
356
Levine
N. D.
(
1988
).
Progress in taxonomy of the Apicomplexan protozoa.
J. Protozool
35
,
518
520
Lingelbach
K.
,
Joiner
K. A.
(
1998
).
The parasitophorous vacuole membrane surrounding Plasmodium and Toxoplasma: an unusual compartment in infected cells.
J. Cell Sci
111
,
1467
1475
Lippincott-Schwartz.
J.
,
Donaldson
J. G.
,
Schweizer
A.
,
Berger
E. G.
,
Hauri
H.-P.
,
Wang
L. C.
,
Klausner
R. D.
(
1990
).
Microtubule-dependent retrograde transport of proteins into the ER in the presence of brefeldin A suggests an ER recycling pathway.
Cell
60
,
821
836
Mellman
I.
,
Simons
K.
(
1992
).
The Golgi complex: in vitroveritas?.
Cell
68
,
829
840
Monro
S.
,
Pelham
H. R. B.
(
1987
).
A C-terminal signal prevents secretion of luminal ER proteins.
Cell
48
,
899
907
Morrissette
N.
,
Bedian
V.
,
Webster
P.
,
Roos
D. S.
(
1994
).
Characterization of extreme apical antigens from Toxoplasma gondii.
Exp. Parasitol
79
,
445
459
Nielsen
H.
,
Engelbrecht
J.
,
Brunak
S.
,
von Heijne
G.
(
1997
).
Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites.
Prot. Engineering
10
,
1
6
Novick
P.
,
Ferro
S.
,
Schekman
R.
(
1981
).
Order of events in the yeast secretory pathway.
Cell
25
,
461
469
Paccaud
J.-P.
,
Reith
W.
,
Carpentier
J.-L.
,
Ravazzoli
M.
,
Amherdt
M.
,
Schekman
R.
,
Orci
L.
(
1996
).
Cloning and functional characterization of mammalian homologues of the COPII components Sec23.
Mol. Biol. Cell
7
,
1535
1546
Palade
G. E.
(
1975
).
Intracellular aspects of the process of protein transport.
Science
189
,
347
358
Pelham
H. R. B.
(
1989
).
Control of protein exit from the endoplasmic reticulum.
Annu. Rev. Cell Biol
5
,
1
23
Rothman
J. E.
,
Orci
L.
(
1992
).
Molecular dissection of the secretory pathway.
Nature
355
,
409
415
Saraste
J.
,
Svensson
K.
(
1991
).
Distribution of the intermediate elements operating in ER to Golgi transport.
J. Cell Sci
100
,
415
430
Schekman
R.
(
1992
).
Genetic and biochemical analysis of vesicular traffic in yeast.
Curr. Opin. Cell Biol
4
,
587
592
Serafini.
T.
,
Stenbeck
G.
,
Brecht
A.
,
Lottspeich
F.
,
Orci
L.
,
Rothman
J. E.
,
Wieland
F. T.
(
1991
).
A coat subunit of Golgi-derived non-clathrin-coated vesicles with homology to the clathrin-coated vesicle coat protein-adaptin.
Nature
349
,
215
220
Shaw
M. K.
,
Tilney
L. G.
(
1992
).
How individual cells develop from a syncytium: merogony in Theileriaparvum (Apicomplexa).
J. Cell Sci
101
,
109
123
Sogin
M. L.
(
1991
).
Early evolution and the origin of eukaryotes.
Curr. Opin. Genet. Dev
1
,
457
463
Stokkermans
T. J. W.
,
Schwartzman
J. D.
,
Keenan
K.
,
Morrissette
N.S.
,
Tilney
L. G.
,
Roos
D. S.
(
1996
).
Inhibition of Toxoplasma gondii replication by dinitroaniline herbicides.
Exp. Parasitol
84
,
355
370
Turner
J. R.
,
Tartakoff
A. M.
(
1989
).
The response of the Golgi complex to microtubule alterations: the roles of metabolic energy and membrane traffic in Golgi complex organization.
J. Cell Biol
109
,
2081
2088
Umebayashi
K.
,
Hirata
A.
,
Fukada
R.
,
Horiuchi
H.
,
Ohta
A.
,
Takagi
M.
(
1997
).
Accumulation of misfolded protein aggregates leads to the formation of Russell body-like dilated endoplasmic reticulum in yeast.
Yeast
13
,
1009
1020
Verkade
P.
,
Simons
K.
(
1997
).
Lipid microdomains and membrane trafficking in mammalian cells.
Histochem. Cell Biol
108
,
211
220
This content is only available via PDF.